Ambion, Inc. proposes to develop an assay platform that will provide rapid quantification of nucleic acids possessing a given sequence from a heterogeneous population of macromolecules. The platform will be especially useful for assessing the viral load of samples too dilute to be accurately quantified using current techniques. The key element of the diagnostic assay is a permeable matrix that contains the enzymes, substrates, and primers necessary to effect the amplification of nucleic acids possessing a specific sequence. Nucleic acids will be extracted from samples and spread over or through the matrix. Amplification will occur at those sites in the matrix where target molecules are located. The matrix will limit the area of amplification of the nucleic adds, thus individual target molecules will give rise to "colonies" of amplified molecules. Nucleic add staining or hybridization to a probe will reveal the colonies, providing an accurate count of the number of amplification competent nucleic acids that were in the samples. Multiple targets could be assayed simultaneously by incorporating multiple primer pairs in the matrix and detecting colonies generated from the different primer pairs with sequence specific probes. PROPOSED COMMERCIAL APPLICATIONS: Greater understanding of the prognostic values of viral, bacterial, and cancer load have created a growing demand for sophisticated diagnostics for quantifying nucleic acid target. The simple design of the proposed diagnostic assay should make it adaptable to a broad spectrum of target nucleic acids, making it possible to quantify virus, pathogenic bacteria, and cancer cells.